The use of polymer materials is expected to considerably improve storage stability and in-vivo particle stability. However, most studies use synthetic polymers produced by emulsion polymerization or the like. Although toxicity is reduced in the synthetic polymers as compared with low-molecular substances, toxicity to some extent should be expected. Therefore, a safer carrier has been demanded.
Natural polymers exhibit high structural stability as with synthetic polymers and have safety much higher than that of synthetic polymers. Thus, the natural polymers have advantages as a carrier. However, a difficult point of the natural polymer carrier as compared with synthetic polymers is a method for producing particles. Spray drying, freeze drying and jet milling can be utilized as methods for producing natural polymer particles. However, in most cases, the particle size is a micron size and is difficult to control.
Comparative survey of Lactoferrin concentration in different milks showed that biggest content of Lactoferrin is in camel milk. Camel milk has 30-100 folds higher concentrations of Lactoferrin than bovine milk. After heat treatment at 85° C. camel milk still contains 37% of Lactoferrin while bovine milk only 1.2% (Conesa C, Sánchez L, Rota C, Pérez M D, Calvo M, Farnaud S, Evans R W (2008), Isolation of lactoferrin from milk of different species: calorimetric and antimicrobial studies, Comp Biochem Physiol B Biochem Mol Biol. 150(1):131-9).
Bovine and camel Lactoferrin are homological with affinity in amount of 75%. Amino acid compound studies showed that camel Lactoferrin is rich with Met while bovine Lactoferrin is rich with Val (Conesa C, Sánchez L, Rota C, Pérez M D, Calvo M, Farnaud S, Evans R W (2008), Isolation of lactoferrin from milk of different species: calorimetric and antimicrobial studies, Comp Biochem Physio B Biochem Mol Biol. 150(1):131-9; Khan J A, Kumar P, Paramasivam M, Yadav R S, Sahani M S, Sharma S, Srinivasan A, Singh T P. (2001), Camel lactoferrin, a transferrin-cum-lactoferrin: crystal structure of camel apolactoferrin at 2.6 A resolution and structural basis of its dual role, J Mol Biol. 309(3):751-61).
Camel Lactoferrin has inhibitory effect on HCV (genotype 4a) higher than human, bovine and sheep lactoferrin (El-Fakharany E M, Sánchez L, Al-Mehdar H A, Redwan E M (2013), Effectiveness of human, camel, bovine and sheep lactoferrin on the hepatitis C virus cellular infectivity: comparison study, Virol J. 10:199. doi: 10.1186/1743-422X-10-199).
Additionally, camel milk lactoferrin was shown to inhibit the proliferation of the colon cancer cell line, HCT-116, in vitro, DNA damage and exhibits antioxidant activities (Habib H M, Ibrahim W H, Schneider-Stock R, Hassan H M. (2013), Camel milk lactoferrin reduces the proliferation of colorectal cancer cells and exerts antioxidant and DNA damage inhibitory activities, Food Chem. 141(1):148-52).
Meanwhile, camel derived Lactoferrin and/or casein are protein insoluble in water contained in camel milk (Farah Z (1993) Composition and characteristics of camel milk, J. Dairy Res. 60(4):603-26).
Since its hydrophobic portion is exposed, Lactoferrin or casein form aggregates under certain conditions. The Lactoferrin and/or casein micelles further gather and form a micelle associate of approximately 100-500 nm. Thus, LF and/or CA micelle has a wide size distribution and is aggregated when placed at acidic pH and supplemented with a sodium or potassium salt.
Glycosaminoglycans (GAGs) are classified into four groups. Heparin/heparan sulfate (HSGAGs) and chondroitin/dermatan sulfate (CSGAGs) are synthesized in the Golgi apparatus, where protein cores made in the rough endoplasmic reticulum are post-translationally modified with O-linked glycosylations by glycosyltransferases forming a proteoglycan. Keratan sulfate may modify core proteins through N-linked glycosylation or O-linked glycosylation of the proteoglycan. The fourth class of GAG, hyaluronic acid, is not synthesized by the Golgi, but rather by integral membrane synthases which immediately secrete the dynamically elongated disaccharide chain.
Lactoferrin (LF), also known as lactotransferrin, is an iron binding glycoprotein with a structure and size that closely resembles (60% sequence homology) to another iron-transporting family, the transferrins (Baker H M, Baker E N (2004), Lactoferrin and iron: structural and dynamic aspects of binding and release, Biometals 17: 209-216.)
Lower concentrations are found in plasma, bile fluids, mucosal secretions, pancreatic fluids and in neutrophils cells (Legrand D, Pierce A, Elass E, Carpentier M, Mariller C, et al. (2008). Lactoferrin structure and functions, Adv Exp Med Biol 606: 163-194.)
Structurally, LF weighs approximately 80 kDa and the polypeptide folds into two globular lobes. Each globe contains two major domains. These domains serve as the binding and glycosylation sites for iron molecules and carbonate ions. Further, depending on the amount of binding, Lf can be classified as apo-LF (iron depleted), monoferric LF (one ferric ion) and holo-Lf (two ferric ions). A possible mechanism for LF's improved binding affinity was suggested to be due to the nature of Lf to be primarily cationic with a high binding affinity to anionic ligands. Therefore this property enables LF to bind to a wide array of “Lf putative receptors” expressed in different cells and organs and aid with the internalization and absorption of LF.
A major problem with currently used cancer treatments like chemotherapy is the inability of drugs to differentiate between malignant and healthy cells leading to severe systemic toxicity. The distinguishing characteristics of NPs such as the small size, large surface-to-volume ratio allowing increased drug encapsulation and ease of functionalizing surface properties to accommodate multiple ligands that can target tumor specific markers is opening new pathways in the search for alternatives to chemotherapy. Thus utilizing natural camel LF and/or casein with or without ionic complex formation with camel derived Glycosmainoglycans (GAGs) and/or inclusion of an active substance as NPs can be a remarkable therapeutic agent, combining LF and/or casein multifunctional properties with the added benefits brought by nanotechnology.